Preclinical Evaluation of Combination Therapy of Rolipram and Minocycline for Arresting Secondary Injury Cascade After Traumatic Brain Injury

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PROJECT SUMMARY For most traumatic brain injuries (TBI), the moment that an impact occurs, there is surprisingly little apparent damage. Yet, within minutes of impact, endogenous responses in the brain amplify and perpetuate the damage manifold. This cascade of responses is called secondary injury, and it is thought that this process can lead to long-term neurological problems. Our goal is to quickly mitigate some of the contributing elements of secondary injury to reduce long-term damage and neurological impairments. We will use a unique combination of drugs, including noninvasive delivery of one drug to the brain using a recently-developed co-polymer, poly (lactide-co-glycolide)-graft-polyethylenimine (PgP) as a nanocarrier. Morphological, physiological, chemical, and behavioral markers of secondary injury will be recorded at several time points up to one month after experimental TBI in mice to determine if the drug combination reduces secondary injury at an earlier time point than either drug alone, and if it maintains this reduction through one month after injury. We will use a unique multimodal system to monitor effects comprised of a newly developed microwire biosensor for glutamate (GLU) and γ-aminobutyric acid (GABA) with a modified micro-prism implant to correlate the dynamics of GLU and GABA signaling and ongoing secondary cellular damage by observing the same cells over time. Our highly sensitive and selective GLU and GABA microbiosensor will be used to evaluate the effect of treatment on reducing excitotoxicity by measuring extracellular concentrations and release dynamics in the cortex, in real time, at 5 time points after TBI versus preinjury baseline. At the same time points, high-resolution multiphoton microscopy through a permanently-implanted, modified prism will be used to facilitate a vertical view of cortical layers 3-6, to monitor development of varicosities (swellings) on dendrites and axons, axonal undulations and retraction bulbs, and the disappearance of axons. Furthermore, by viewing the same cells over time, we will compare the ability of the drugs to resolve dendritic and axonal varicosities on cells exhibiting damage in previous imaging sessions. Therapeutic effects will also be evaluated using standard behavioral tests for motor coordination, exploration, and memory, and by using established assays and antibody staining and cytokine assays for molecular biomarkers of secondary injury. By comparing the results from our novel combination of optical imaging and real-time glutamate and GABA signaling with the results from well-established behavioral and molecular biomarker tests at matched time points, we will demonstrate the utility of this longitudinal, optical- biosensor system to quantify secondary damage and its resolution. Notably, this longitudinal approach will require fewer animals because each animal serves as its own control, reducing variability, and each is used at multiple time points. In addition, optical-biosensor data from injured, vehicle-treated mice will provide new insights for better understanding the cascade of secondary injury and neural degeneration after TBI. Furthermore, this new optical-biosensor system will become a valuable tool for evaluating other drugs and for optimizing therapeutic windows.